Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line
- Autores
- Bermúdez, Vicente; Asatryan, Asatryan A.; Mukherjee, P. K.; Giusto, Norma Maria; Bazan, Nicolás Guillermo; Mateos, Melina Valeria
- Año de publicación
- 2021
- Idioma
- inglés
- Tipo de recurso
- documento de conferencia
- Estado
- versión publicada
- Descripción
- The retinal pigment epithelium (RPE) plays critical roles in the correct function of the neural retina and photoreceptor survival. Classical phospholipases D (PLD1 and 2) hydrolyze phosphatidylcholine (PC) to generate phosphatidic acid (PA) and choline. PA can be further dephosphorylated to diacylglycerol (DAG) by lipid phosphate phosphatases (LPPs). DAG and PA, as bioactive lipids, can modulate the activity of various proteins involved in cell signaling events, such as protein kinases C and the mTOR (mammalian target of rapamycin) complex, among others. Our previous studies demonstrated for the first time the participation of classical PLDs in the inflammatory response and in the autophagic process of RPE (ARPE-19 and D407) cells exposed to lipopolysaccharide (LPS). The aim of the present work was to study PLD1 and PLD2 expression in a new human RPE cell line (ABC cells), spontaneously arisen from a primary RPE cell culture. Western blot assays (WB) show that both classical PLDs are expressed in ABC cells. Using PLD1 and PLD2 siRNA, we were able to partially decrease the expression of PLD1 (by 42%) and PLD2 (by30 %). Since PLD-generated PA activates mTORC1, the main inhibitor of autophagy initiation, we wanted to study the effect of classical PLDs silencing on mTOR activation. For this purpose, WB assays were performed in order to study the mTOR downstream effector S6 kinase (S6K) activation (phosphorylation) in ABC cells transfected with PLD1 and PLD2 siRNA. Our results show that in ABC cells transfected with PLD1 and PLD2 siRNA S6K activation is reduced by 34%. This result is in accordance with the increased autophagic process induced by PLD1 and PLD2 pharmacological inhibitors, previously observed in D407 RPE cells. Since it was previously demonstrated that the PLD pathway can modulate the phagocytic process in macrophages, we wanted to evaluate the effect of PLD1 and PLD2 silencing on the photoreceptor outer segment (POS) phagocytic process in ABC cells. With this aim, ABC cells were incubated with POS for 16 h, and total (bound + internal) and internalized POS were measured by WB using an anti-rhodopsin antibody. Under basal conditions, PLD1 and PLD2 silencing seem not to significantly affect POS phagocytosis by ABC cells. In conclusion, our results demonstrate the expression of classical PLD isoforms in a new RPE cell line and their role in the modulation of the mTOR/S6K pathway. Further experiments are needed to fully elucidate the role of classical PLDs in the phagocytic process of ABC RPE cells exposed to inflammatory conditions. The results presented herein, together with our previous findings, contribute to the knowledge of the molecular basis of retinal inflammatory and degenerative diseases, such as diabetic retinopathy, aged-related macular degeneration and endophthalmitis, among others.
Fil: Bermúdez, Vicente. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina
Fil: Asatryan, Asatryan A.. Louisiana State University Health Sciences Center New Orleans (lsu Health);
Fil: Mukherjee, P. K.. Louisiana State University Health Sciences Center New Orleans (lsu Health);
Fil: Giusto, Norma Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina
Fil: Bazan, Nicolás Guillermo. Louisiana State University Health Sciences Center New Orleans (lsu Health);
Fil: Mateos, Melina Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina
LVI Annual Meeting Argentine Society for Biochemistry and Molecular Biology and XV Annual Meeting Argentinean Society for General Microbiology
Argentina
Sociedad Argentina de Investigación en Bioquímica y Biología Molecular
Sociedad Argentina de Microbiología General - Materia
-
PHOSPHOLIPASE D
PHAGOCYTOSIS
RETINAL PIGMENT EPITHELIUM - Nivel de accesibilidad
- acceso abierto
- Condiciones de uso
- https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
- Repositorio
- Institución
- Consejo Nacional de Investigaciones Científicas y Técnicas
- OAI Identificador
- oai:ri.conicet.gov.ar:11336/229410
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Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell lineBermúdez, VicenteAsatryan, Asatryan A.Mukherjee, P. K.Giusto, Norma MariaBazan, Nicolás GuillermoMateos, Melina ValeriaPHOSPHOLIPASE DPHAGOCYTOSISRETINAL PIGMENT EPITHELIUMhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1The retinal pigment epithelium (RPE) plays critical roles in the correct function of the neural retina and photoreceptor survival. Classical phospholipases D (PLD1 and 2) hydrolyze phosphatidylcholine (PC) to generate phosphatidic acid (PA) and choline. PA can be further dephosphorylated to diacylglycerol (DAG) by lipid phosphate phosphatases (LPPs). DAG and PA, as bioactive lipids, can modulate the activity of various proteins involved in cell signaling events, such as protein kinases C and the mTOR (mammalian target of rapamycin) complex, among others. Our previous studies demonstrated for the first time the participation of classical PLDs in the inflammatory response and in the autophagic process of RPE (ARPE-19 and D407) cells exposed to lipopolysaccharide (LPS). The aim of the present work was to study PLD1 and PLD2 expression in a new human RPE cell line (ABC cells), spontaneously arisen from a primary RPE cell culture. Western blot assays (WB) show that both classical PLDs are expressed in ABC cells. Using PLD1 and PLD2 siRNA, we were able to partially decrease the expression of PLD1 (by 42%) and PLD2 (by30 %). Since PLD-generated PA activates mTORC1, the main inhibitor of autophagy initiation, we wanted to study the effect of classical PLDs silencing on mTOR activation. For this purpose, WB assays were performed in order to study the mTOR downstream effector S6 kinase (S6K) activation (phosphorylation) in ABC cells transfected with PLD1 and PLD2 siRNA. Our results show that in ABC cells transfected with PLD1 and PLD2 siRNA S6K activation is reduced by 34%. This result is in accordance with the increased autophagic process induced by PLD1 and PLD2 pharmacological inhibitors, previously observed in D407 RPE cells. Since it was previously demonstrated that the PLD pathway can modulate the phagocytic process in macrophages, we wanted to evaluate the effect of PLD1 and PLD2 silencing on the photoreceptor outer segment (POS) phagocytic process in ABC cells. With this aim, ABC cells were incubated with POS for 16 h, and total (bound + internal) and internalized POS were measured by WB using an anti-rhodopsin antibody. Under basal conditions, PLD1 and PLD2 silencing seem not to significantly affect POS phagocytosis by ABC cells. In conclusion, our results demonstrate the expression of classical PLD isoforms in a new RPE cell line and their role in the modulation of the mTOR/S6K pathway. Further experiments are needed to fully elucidate the role of classical PLDs in the phagocytic process of ABC RPE cells exposed to inflammatory conditions. The results presented herein, together with our previous findings, contribute to the knowledge of the molecular basis of retinal inflammatory and degenerative diseases, such as diabetic retinopathy, aged-related macular degeneration and endophthalmitis, among others.Fil: Bermúdez, Vicente. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; ArgentinaFil: Asatryan, Asatryan A.. Louisiana State University Health Sciences Center New Orleans (lsu Health);Fil: Mukherjee, P. K.. Louisiana State University Health Sciences Center New Orleans (lsu Health);Fil: Giusto, Norma Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; ArgentinaFil: Bazan, Nicolás Guillermo. Louisiana State University Health Sciences Center New Orleans (lsu Health);Fil: Mateos, Melina Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; ArgentinaLVI Annual Meeting Argentine Society for Biochemistry and Molecular Biology and XV Annual Meeting Argentinean Society for General MicrobiologyArgentinaSociedad Argentina de Investigación en Bioquímica y Biología MolecularSociedad Argentina de Microbiología GeneralTech Science Press2021info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectReuniónJournalhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/vnd.openxmlformats-officedocument.wordprocessingml.documentapplication/pdfhttp://hdl.handle.net/11336/229410Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line; LVI Annual Meeting Argentine Society for Biochemistry and Molecular Biology and XV Annual Meeting Argentinean Society for General Microbiology; Argentina; 2020; 74-750327-9545CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.techscience.com/biocell/v45nSuppl.1/42376/pdfNacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:11:17Zoai:ri.conicet.gov.ar:11336/229410instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 10:11:17.911CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse |
dc.title.none.fl_str_mv |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
title |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
spellingShingle |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line Bermúdez, Vicente PHOSPHOLIPASE D PHAGOCYTOSIS RETINAL PIGMENT EPITHELIUM |
title_short |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
title_full |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
title_fullStr |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
title_full_unstemmed |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
title_sort |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line |
dc.creator.none.fl_str_mv |
Bermúdez, Vicente Asatryan, Asatryan A. Mukherjee, P. K. Giusto, Norma Maria Bazan, Nicolás Guillermo Mateos, Melina Valeria |
author |
Bermúdez, Vicente |
author_facet |
Bermúdez, Vicente Asatryan, Asatryan A. Mukherjee, P. K. Giusto, Norma Maria Bazan, Nicolás Guillermo Mateos, Melina Valeria |
author_role |
author |
author2 |
Asatryan, Asatryan A. Mukherjee, P. K. Giusto, Norma Maria Bazan, Nicolás Guillermo Mateos, Melina Valeria |
author2_role |
author author author author author |
dc.subject.none.fl_str_mv |
PHOSPHOLIPASE D PHAGOCYTOSIS RETINAL PIGMENT EPITHELIUM |
topic |
PHOSPHOLIPASE D PHAGOCYTOSIS RETINAL PIGMENT EPITHELIUM |
purl_subject.fl_str_mv |
https://purl.org/becyt/ford/1.6 https://purl.org/becyt/ford/1 |
dc.description.none.fl_txt_mv |
The retinal pigment epithelium (RPE) plays critical roles in the correct function of the neural retina and photoreceptor survival. Classical phospholipases D (PLD1 and 2) hydrolyze phosphatidylcholine (PC) to generate phosphatidic acid (PA) and choline. PA can be further dephosphorylated to diacylglycerol (DAG) by lipid phosphate phosphatases (LPPs). DAG and PA, as bioactive lipids, can modulate the activity of various proteins involved in cell signaling events, such as protein kinases C and the mTOR (mammalian target of rapamycin) complex, among others. Our previous studies demonstrated for the first time the participation of classical PLDs in the inflammatory response and in the autophagic process of RPE (ARPE-19 and D407) cells exposed to lipopolysaccharide (LPS). The aim of the present work was to study PLD1 and PLD2 expression in a new human RPE cell line (ABC cells), spontaneously arisen from a primary RPE cell culture. Western blot assays (WB) show that both classical PLDs are expressed in ABC cells. Using PLD1 and PLD2 siRNA, we were able to partially decrease the expression of PLD1 (by 42%) and PLD2 (by30 %). Since PLD-generated PA activates mTORC1, the main inhibitor of autophagy initiation, we wanted to study the effect of classical PLDs silencing on mTOR activation. For this purpose, WB assays were performed in order to study the mTOR downstream effector S6 kinase (S6K) activation (phosphorylation) in ABC cells transfected with PLD1 and PLD2 siRNA. Our results show that in ABC cells transfected with PLD1 and PLD2 siRNA S6K activation is reduced by 34%. This result is in accordance with the increased autophagic process induced by PLD1 and PLD2 pharmacological inhibitors, previously observed in D407 RPE cells. Since it was previously demonstrated that the PLD pathway can modulate the phagocytic process in macrophages, we wanted to evaluate the effect of PLD1 and PLD2 silencing on the photoreceptor outer segment (POS) phagocytic process in ABC cells. With this aim, ABC cells were incubated with POS for 16 h, and total (bound + internal) and internalized POS were measured by WB using an anti-rhodopsin antibody. Under basal conditions, PLD1 and PLD2 silencing seem not to significantly affect POS phagocytosis by ABC cells. In conclusion, our results demonstrate the expression of classical PLD isoforms in a new RPE cell line and their role in the modulation of the mTOR/S6K pathway. Further experiments are needed to fully elucidate the role of classical PLDs in the phagocytic process of ABC RPE cells exposed to inflammatory conditions. The results presented herein, together with our previous findings, contribute to the knowledge of the molecular basis of retinal inflammatory and degenerative diseases, such as diabetic retinopathy, aged-related macular degeneration and endophthalmitis, among others. Fil: Bermúdez, Vicente. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina Fil: Asatryan, Asatryan A.. Louisiana State University Health Sciences Center New Orleans (lsu Health); Fil: Mukherjee, P. K.. Louisiana State University Health Sciences Center New Orleans (lsu Health); Fil: Giusto, Norma Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina Fil: Bazan, Nicolás Guillermo. Louisiana State University Health Sciences Center New Orleans (lsu Health); Fil: Mateos, Melina Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina LVI Annual Meeting Argentine Society for Biochemistry and Molecular Biology and XV Annual Meeting Argentinean Society for General Microbiology Argentina Sociedad Argentina de Investigación en Bioquímica y Biología Molecular Sociedad Argentina de Microbiología General |
description |
The retinal pigment epithelium (RPE) plays critical roles in the correct function of the neural retina and photoreceptor survival. Classical phospholipases D (PLD1 and 2) hydrolyze phosphatidylcholine (PC) to generate phosphatidic acid (PA) and choline. PA can be further dephosphorylated to diacylglycerol (DAG) by lipid phosphate phosphatases (LPPs). DAG and PA, as bioactive lipids, can modulate the activity of various proteins involved in cell signaling events, such as protein kinases C and the mTOR (mammalian target of rapamycin) complex, among others. Our previous studies demonstrated for the first time the participation of classical PLDs in the inflammatory response and in the autophagic process of RPE (ARPE-19 and D407) cells exposed to lipopolysaccharide (LPS). The aim of the present work was to study PLD1 and PLD2 expression in a new human RPE cell line (ABC cells), spontaneously arisen from a primary RPE cell culture. Western blot assays (WB) show that both classical PLDs are expressed in ABC cells. Using PLD1 and PLD2 siRNA, we were able to partially decrease the expression of PLD1 (by 42%) and PLD2 (by30 %). Since PLD-generated PA activates mTORC1, the main inhibitor of autophagy initiation, we wanted to study the effect of classical PLDs silencing on mTOR activation. For this purpose, WB assays were performed in order to study the mTOR downstream effector S6 kinase (S6K) activation (phosphorylation) in ABC cells transfected with PLD1 and PLD2 siRNA. Our results show that in ABC cells transfected with PLD1 and PLD2 siRNA S6K activation is reduced by 34%. This result is in accordance with the increased autophagic process induced by PLD1 and PLD2 pharmacological inhibitors, previously observed in D407 RPE cells. Since it was previously demonstrated that the PLD pathway can modulate the phagocytic process in macrophages, we wanted to evaluate the effect of PLD1 and PLD2 silencing on the photoreceptor outer segment (POS) phagocytic process in ABC cells. With this aim, ABC cells were incubated with POS for 16 h, and total (bound + internal) and internalized POS were measured by WB using an anti-rhodopsin antibody. Under basal conditions, PLD1 and PLD2 silencing seem not to significantly affect POS phagocytosis by ABC cells. In conclusion, our results demonstrate the expression of classical PLD isoforms in a new RPE cell line and their role in the modulation of the mTOR/S6K pathway. Further experiments are needed to fully elucidate the role of classical PLDs in the phagocytic process of ABC RPE cells exposed to inflammatory conditions. The results presented herein, together with our previous findings, contribute to the knowledge of the molecular basis of retinal inflammatory and degenerative diseases, such as diabetic retinopathy, aged-related macular degeneration and endophthalmitis, among others. |
publishDate |
2021 |
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2021 |
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http://hdl.handle.net/11336/229410 Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line; LVI Annual Meeting Argentine Society for Biochemistry and Molecular Biology and XV Annual Meeting Argentinean Society for General Microbiology; Argentina; 2020; 74-75 0327-9545 CONICET Digital CONICET |
url |
http://hdl.handle.net/11336/229410 |
identifier_str_mv |
Phospholipase D (PLD) 1 and 2 expression in ABC cells, a new retinal pigment epithelium cell line; LVI Annual Meeting Argentine Society for Biochemistry and Molecular Biology and XV Annual Meeting Argentinean Society for General Microbiology; Argentina; 2020; 74-75 0327-9545 CONICET Digital CONICET |
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eng |
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eng |
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Tech Science Press |
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